Specific cancer cell targeted RNAi drug delivery is a very promising strategy for many disease treatments including cancer. Exosomes, naturally derived nano vesicles from the endosome membrane of cells, showed very encouraging ability to deliver siRNA into cells in vitro. But to conquer the physiological barriers and achieve therapeutic effect in vivo, exosomes with specific cancer cell targeting property are demanded. Disclosed herein are methods and compositions for displaying ligands onto exosome surface post-biogenesis. RNA nanostructures can be utilized as a tool to display the RNA or chemical based ligand onto exosome surface, thus increase their cell targeting specificity and thus can be used for specific delivery of therapeutic reagent, such as RNAi therapeutics, to the targeted cells.
RNA nanostructures derived from packaging RNA of phi29 DNA packaging motor have shown great promise for drug delivery. The 3WJ domain of pRNA is highly thermodynamically stable, can be formed from 3 pieces of short RNA oligonucleotides with high affinity. Furthermore, when using the 3WJ as a core for building RNA nanoparticles, it can drive the global folding of the RNA nanoparticle and ensure the correct folding of fused aptamer sequences to remain functional. Cholesterol was applied to modified pRNA-3WJ for displaying ligand onto exosome surface. The results showed that both chemical ligand and RNA aptamer can be displayed on exosome via cholesterol modified pRNA 3WJ. Ligand displaying exosomes have enhanced specific tumor binding efficiency in vitro. In the animal experiment, ligand displaying exosomes showed specific accumulation in tumor after systemic injection. Exosome was further loaded with siRNA, ligand displaying exosomes can enhance the siRNA delivery efficiency to target cancer cells in vitro and in vivo.
RNAi therapeutics is very promising for treating various diseases including cancer, since it has the ability to modify disease gene expression. However, despite years of extensive research, an efficient and biocompatible RNAi delivery system is still lacking. Though liposomes show great success for siRNA delivery in vitro, but when systemically administering in vivo, the problems persist of liver accumulation and freeze-thaw cycles causing instability in the final product.
Exosomes, which are nano-scaled vesicles originated from cell endosome membrane, have been studied extensively as RNAi drug delivery system recently. But to achieve specific cancer cell targeting is still challenging. Current technologies are exploring expressing cancer cell specific ligand on exosome generating cells to increase the exosome specificity, such as overexpression peptide ligands on the exosome membrane as fusion protein on HEK293T cells. But one problem for using fusion peptides for targeted exosome delivery is that the displayed peptide can be degraded during exosome biogenesis.
What is needed in the art is RNA ligand-displaying exosomes for specific delivery of therapeutics to cells by RNA technology.